High-throughput screening (HTS) is a method used in life science research and the biopharmaceutical industry for drug discovery, toxicology testing, and functional genomics. Typically, HTS is used to rapidly determine the physiological response of groups of cells to various combinations and quantities of biologically active chemical compounds and biomaterials surrounding the cell.
Cellular activity is also influenced by applied mechanical stimulation, which has been shown to have a strong impact on biological function in certain types of cells (McBeath, et al., Dev. Cell 2004; Wang & Thampatty, Biomech Model Mechanobiol 2006; Saha et al., J Cell Physiol, 2006). Existing experimental techniques are unable to adequately characterize cellular response to varying degrees of mechanical stimulation with a high accuracy in a high-throughput manner. These limitations have prevented systematic investigations into the effects of mechanical stimuli on cell behaviour and hindered discovery of new control strategies for cell-based therapies.
Furthermore, despite the demonstrated individual importance of mechanical forces; chemical cues; and the composition and structure of surrounding biomaterials in regulated cellular function, the lack of HTS techniques for mechanical factors precludes the ability to effectively study combinations of these various parameters. This patent application discloses a system designed to meet this need for rapidly probing either single cells or colonies of cells.
Existing low-throughput experimental techniques in this field make use of three main mechanical loading schemes to probe cellular response: compressive loading, deformation of the substrate to which cells adhere, and fluid flow-induced shear stresses. U.S. Pat. No. 6,048,723 discloses a flexible bottom culture plate for applying mechanical loads to cell culture; U.S. Pat. No. 6,218,178 discloses the loading assembly for the plates of U.S. Pat. No. 6,048,723; U.S. Pat. No. 6,645,759 discloses a device for growing cells in culture under shear stress and/or strain; and U.S. Pat. No. 6,037,141 discloses a system for culturing cells under compression conditions. However, the systems described the cited US patents are all low-throughput, applying a single strain across at most, six experimental locations. This drawback significantly impacts the time required to perform such studies. It also precludes the ability to perform combinatorial manipulation of chemical and mechanical parameters, as can be performed in our disclosed invention.
Moreover, there are two modes of cell culture: two-dimensional culture on a flat surface, and three-dimensional culture within a porous biomaterial. Each of these culture techniques and loading scenarios provide insight into the inner workings of the cell, but typically require radically different experimental setups.
Microsystems are engineered systems with critical structural or functional features of micrometers, where the microfabricated component of the system typically ranges in size from millimeters to centimeters. They have such advantages as low cost, small size, minimal reagent consumption and fast response time. Because of the reduced system footprint, a dense array of functional sub-units is possible, and as such they are ideal for developing array based HTS systems. Similarities between system feature sizes and the size of a cell make this technology suitable for developing HTS systems for single- or multi-cell biology. Advances in microfabrication have enabled the rapid development of complex, elastomeric, monolithic polymer structures with well-defined features with a resolution of micrometers. To provide an example, these techniques—termed Multilayer Soft Lithography (MSL)—have been used to develop a fully controllable microfluidic network, actuated by a number of 2-state valves (Unger et al., “Monolithic microfabricated valves and pumps by multilayer soft lithography,” Science, vol. 288, pp. 113-6, Apr. 7 2000; and U.S. Pat. No. 6,793,753).
In view of the foregoing, an improved apparatus, system and method for HTS applications is desirable.
The disclosed invention introduces new aspects in MSL device development, including the use of mechanical solid elements in an all-polymer device, and the application of a single pressure load to obtain a range of mechanical activity.